How DNA methylation patterns are shaped during development - how methylation and demethylation are targeted to specific loci at specific times - is not known. Germ cells establish methylation imprints depending on the sex of the gonad after erasing the male and female -specific marks inherited from the previous generation. Our main hypothesis is that DNA cytosine 5-hydroxymethylation and chromatin composition are important components of the imprint cycle, being involved in the erasure and establishment phases in germ cells and in the maintenance phase in the soma. To demonstrate this, we will analyze DNA methylation, DNA cytosine hydroxymethylation and chromatin globally and at specific imprinted genes in germ cells and somatic cells. To test the role of specific epigenetic modifiers in the cycle of imprinting, we will genetically perturb these functions during the erasure, establishment and maintenance phases of the imprinting cycle. These results collectively will help understand how DNA methylation, DNA hydroxymethylation and chromatin marks collaborate in the epigenetic remodeling of germ cells and in erasing, establishing and maintaining genomic imprints. Correct global epigenetic remodeling and imprint reseting during embryonic-fetal germ cell development is essential for the health of the next generation and for future generations. PUBLIC HEALTH RELEVANCE: We propose to test the role of DNA cytosine hydroxymethylation and chromatin in the erasure, establishment and maintenance of genomic imprints. Experiments using genome-wide mapping methods, novel allele- specific assays and mouse genetic tools will collectively help understand how DNA methylation, DNA hydroxymethylation and chromatin marks collaborate in the global epigenetic remodeling of germ cells and in erasing, establishing and maintaining genomic imprints. Correct global epigenetic remodeling and imprint resetting during embryonic-fetal germ cell development is essential for the health of the next generation and for future generations.